Project Summary: Glucocorticoids are indispensable in the treatment of inflammation although they are not effective in treating neutrophilic lung inflammation in severe asthma and acute respiratory distress syndrome. Our long-term goal is to develop new glucocorticoid regimens with improved efficacy/risk ratios. The goal of this proposal is to determine whether eliminating G-CSF signaling will allow glucocorticoids to promote neutrophil apoptosis and ameliorate lung inflammation. We have recently discovered that glucocorticoids significantly elevate G-CSF, a neutrophil promoting cytokine. G-CSF in recent years has been recognized as a cause of steroid-resistant lung inflammatory disorders. We hypothesize that glucocorticoids protect neutrophil from apoptosis and enhance neutrophil function via stimulation of G-CSF. G-CSF induction is initiated by GR and NF-?B p50 interaction on the G-CSF promoter. Glucocorticoid-induction of G-CSF production in lung resident cells sustains lung inflammation. Blocking G-CSF signaling will increase the efficacy of glucocorticoids in circumventing neutrophil-driven lung inflammation. To test our hypotheses, we will examine systems in vitro and in vivo. Studies in Aim 1 will test the hypothesis that GR and p50 interactions mediate G-CSF expression in various cell types in lungs. Studies in Aim 2 will determine the role of glucocorticoid-induced G-CSF in neutrophil survival, functions, trafficking, and neutrophilic lung inflammation in two murine models. We will use three approaches to block G-CSF induction: using G-CSFR null mice, anti-G-CSF neutralizing antibodies, and two natural NF-?B inhibitors that we have identified to be able to inhibit G-CSF-induction: curcumin and parthenolide. Our preliminary data indicate that both the glucocorticoid receptor (GR) and NF-?B were required for G-CSF induction and p50, but not p65, was recruited to the same region of G-CSF promoter as GR. Multiple lung resident cells and infiltrating leukocytes produced G-CSF upon glucocorticoid stimulation. Blocking G-CSF signaling using neutralizing antibodies abolished glucocorticoid and IL-1? protection of neutrophils from undergoing apoptosis. Curcumin in combination with glucocorticoids reduced lung G-CSF and neutrophil numbers in an LPS lung injury model. The proposed studies will improve our understanding of the role of glucocorticoid-induced G-CSF in neutrophil-dominant glucocorticoid-resistant diseases. Inhibiting G- CSF signaling is anticipated to increase the ability of glucocorticoids to suppress the airway-damaging activities of neutrophils and ameliorate lung inflammation.